Sialic acid is an N- or O-substituted derivative of neuraminic acid. The N-substituted versions generally bear either an acetyl or a glycolyl group. In contrast, the O-substituted hydroxyl group may vary considerably, e.g., acetyl, lactyl, methyl, sulfate and phosphate groups. Polysialic acids are also quite common in which N-acetyl neuraminic acid residues are linked via the C2 ketal OH to another molecule by a glycosidic bond, e.g., poly alpha (2→8) N-acetyl neuraminic acid.
The sialic acids are biologically important carbohydrates found in organisms ranging from bacteria to humans. They are common features decorating the terminal ends of glycoproteins, glycans and glycosphingolipids, as well as other molecules. They mediate myriad normal cellular activities. This includes stabilizing glycoconjugates in cell membranes, regulating cell-cell interactions, acting as chemical messengers, regulating transmembrane receptor function, affecting membrane transport, controlling the half-lives of circulating glycoproteins and cells, and contributing to the permselectivity of the glomerular endothelium. See for review: Angata and Varki Chem. Rev. (2002) 102:439.
Given their prominent role in normal cellular activity, sialic acid and its derivatives have been used as markers for abnormal cellular processes such as cancer. (O'Kennedy et al., Cancer Lett., 1991 58:91; Vedralova et al. Cancer Lett. 1994 78:171; and Horgan et al., Clin. Chim. Acta., 1982 118:327; and Narayanan, S. Ann. Clin. Lab. Sci. 1994 24:376). For instance, cancer cells that can metastasize often have larger amounts of sialic acid-modified glycoproteins, which may help them enter the blood stream. Also, the sialic acid of tumor cells is modified in ways that differ from normal cells (Hakamori Cancer Res. 1996, 56:5309, Dall'Olio Clin. Mol. Pathol. 1996, 49:M126, Kim and Varki Glycoconj. J. 1997, 14:569).
One sialic acid derivative thought to be uncommon in normal cells, but present on cancer cells is de-N-acetyl sialic acid (Hanai et al J. Biol. Chem. 1988, 263:6296, Manzi et al J. Biol. Chem. 1990, 265:1309, Sjoberg et al J. Biol. Chem. 1995, 270:2921, Chamas et al 1999, Cancer Res. 59:1337; and Popa et al Glycobiology. 2007 17:367).
SEAM 3 is a murine monoclonal antibody that binds to poly alpha (2→8) N-acetyl neuraminic acid (polysialic acid or PSA) that contains de-N-acetyl residues (i.e., neuraminic acid) (Moe et al, Infect. Immun., 2005, 73:2123). SEAM 3 mediates bacteriolysis of Neisseria meningitidis group B (NmB) bacteria in the presence of exogenous complement and provides passive protection in an in vivo infant rat model of meningococcal bacteremia. SEAM 3 also binds to PSA antigens expressed in a variety of human tumors resulting in arrest of cell growth and a reduction in viability by inducing apoptosis and cell death.
Sodium borohydride is a reducing agent used to reduce aldehydes, ketones, imines, acid chlorides, and anhydrides. However, under mild basic conditions, it has also been shown to de-N-acetylate glycosaminoglycuronans (Hirano et al, Connect Tissue Res, 1975, 3:73) and cleave amide bonds in polypeptides (Shimamura et al, Arch Biochem Biophys, 1984, 232:699). A product of the sodium borohydride reduction is sodium borate. Borates and boranes are known to form cyclic esters with a variety of 1,2 or 1,3-diols in a 1:1 or 1:2 ratio. At pH<9 borates form complexes with the alpha caroboxylate of N-acetyl neuraminic acid and at pH >9 with the glycerol moiety at the non-reducing end (Djanashvili et al, Chem Eur J, 2005, 11:4010).
Relevant Literature
Amino sugars, derivatives and related literature of interest are reported in the following U.S. Pat. Nos.: 4,021,542; 4,062,950; 4,175,123; 4,216,208; 4,254,256; 4,314,999; 4,656,159; 4,713,374; 4,797,477; 4,803,303; 4,840,941; 4,914,195; 4,968,786; 4,983,725; 5,231,177; 5,243,035; 5,264,424; 5,272,138; 5,332,756; 5,667,285; 5,674,988; 5,759,823; 5,962,434; 6,075,134; 6,110,897; 6,274,568; 6,407,072; 6,458,937; 6,548,476; 6,697,251; 6,680,054; 6,936,701; and 7,070,801, and in the following references: Angata and Varki Chem. Rev. 2002, 102:439; Hakamori Cancer Res. 1996, 56:5309; Dall'Olio Clin. Mol. Pathol. 1996, 49:M126; Kim and Varki Glycoconj. J. 1997, 14:569; Hanai et al J. Biol. Chem. 1988, 263:6296; Manzi et al J. Biol. Chem. 1990, 265:1309; Sjoberg et al J. Biol. Chem. 1995, 270:2921; Chamas et al Cancer Res. 1999, 59:1337; Popa et al Glycobiology. 2007 17:367; Kayser et al J. Biol. Chem. 1992 267:16934; Keppler et al Glycobiology 2001, 11:1R; Luchansky et al Meth. Enzymol. 2003, 362:249; Oetke et al Eur. J. Biochem. 2001, 268:4553; Collins et al Glycobiology 2000, 10:11; and Bardor et al J. Biol. Chem. 2005, 280:4228.
The antibody SEAM 3 is reported in Moe et al, Infect. Immun., 2005, 73:2123. Sodium borohydride reactions and related are reported in various references, such as Hirano et al, Connect Tissue Res, 1975, 3:73; Shimamura et al, Arch Biochem Biophys, 1984, 232:699; and Djanashvili et al, Chem Eur J, 2005, 11:4010. See also See also US 2007/0010482; U.S. application Ser. No. 11/645,255, filed Dec. 22, 2006; WO 2006/002402; and PCT application serial no. PCT/US2006/04885, filed Dec. 22, 2006.